4.14.2. Antimicrobial Activity of Active Films Application on Cheese against *S. aureus*

The antimicrobial activity of active films on toxigenic strain *S. aureus* ATCC1538 growth in cottage cheese was studied considering the possible diffusion of essential oil from the film matrix into the soft cheese. The cottage cheese divided into 5 treatments consisting of (1) control uncoated sample (C); (2) sample coated with ALG/G film without the addition of the hybrid nanostructure active hybrid nanostructure TO@NZ; (3) sample coated with ALG/G/5TO@NZ film; 4) sample coated with ALG/G/10 TO@NZ films; and (5) sample coated with ALG/G/15TO@NZ. Cottage cheese for each treatment was weighed into 50±0.5 g portions and placed in a stomacher bag. The cheese was inoculated with a fresh suspension of pathogenic *S. aureus* ATCC1538 strain in a final count of 10<sup>4</sup> cfu/g. The cheese was homogenized for 2 min by the stomacher (Seward 400; UK) and placed into sterile plastic petri-dish (9 cm diameter) as a thin layer. The film materials prepared as described above were applied to the upper surfaces of the cheese except for control (C); and the plates after being covered with their lid; were held at 10 ◦C for 10 days and sampled at days 0; 2; 5; 7; 10. The choice of time and temperature abuse of samples was selected as the most common conditions of unproperly preserved foodstuffs in the retail market; causing foodborne illness. At each sampling interval, duplicate plates from each treatment were aseptically opened and a 10 g-portion was weighed in a Stomacher-bag and homogenized in sterile peptone salt solution (Merck; Germany) in a stomacher (Seward 400; UK) for 1 min to make the initial dilution (10−<sup>1</sup> ). Appropriate serial dilutions (10−1–10−<sup>5</sup> ) were spread plated on Baird Parker agar with egg yolk tellurite emulsion (BPA; Merck; Germany) and incubated at 37 ◦C for 48 h for *Staphylococcus aureus* ATCC1538 counts.
